Bifidobacterium animalis subsp. Lactis KW365 capable of fermenting medicinal and edible food materials to reduce uric acid and application of bifidobacterium animalis subsp. Lactis KW365

Abstract

The invention discloses bifidobacterium animalis subsp. Lactis KW365 capable of fermenting medicinal and edible food materials and reducing uric acid and application of the bifidobacterium animalis subsp. Lactis KW365. The bifidobacterium animalis subsp. Lactis KW365 is preserved in the China Center for Type Culture Collection, the preservation date is May 28, 2024, and the preservation number is CCTCC NO: M 20241082. The bifidobacterium animalis subsp. Lactis KW365 has good tolerance to Chinese herbal medicines, and the bifidobacterium animalis subsp. Lactis KW365 and fermentation products prepared by fermenting specific Chinese herbal medicines by using the bifidobacterium animalis subsp. Lactis KW365 have remarkable antibacterial effect and virus killing effect. Besides, KW365 is used for fermenting specific medicinal and edible food materials, the strain and a fermentation product can effectively reduce the serum uric acid level and relieve hyperuricemia and have a protection effect on kidney injury, particularly, the fermentation product effect is remarkable and better, and a remarkable synergistic interaction effect is achieved between the KW365 and specific Chinese herbal medicines.

Description

Technical Field

[0001] This invention belongs to the technical field of Bifidobacterium lactis subspecies, and particularly relates to a strain of Bifidobacterium lactis KW365 that can ferment food ingredients that are both food and medicine to lower uric acid and its applications. Background Technology

[0002] Hyperuricemia is a metabolic disease caused by purine metabolism disorder, resulting in a persistently elevated blood uric acid concentration. Normally, the body can metabolize purines from food (commonly found in animal organs, seafood, and other natural components), but patients often have a reduced ability to process purines, leading to uric acid buildup in the blood. These patients often also have obesity, abnormal blood sugar (type 2 diabetes), dyslipidemia, and high blood pressure. These health risks can easily exacerbate each other, a condition medically termed metabolic syndrome. Hyperuricemia includes primary and secondary hyperuricemia. Primary hyperuricemia is mainly caused by congenital purine metabolism abnormalities. These patients often have multiple health problems such as obesity, diabetes, dyslipidemia, hypertension, and arteriosclerosis; this coexistence of multiple conditions is commonly referred to as metabolic syndrome. Secondary hyperuricemia is usually caused by other diseases or medications. Common causes include hematologic disorders (such as leukemia and multiple myeloma), chronic kidney disease, and long-term use of certain medications (such as the diuretic furosemide and the antipyretic analgesic aspirin).

[0003] Traditional Chinese medicine extracts refer to the effective components or parts extracted from medicinal materials such as plants and minerals through physical or chemical methods, and then processed into intermediate products through concentration and purification. They serve as raw materials or excipients for traditional Chinese medicine preparations, health products, and food additives. Compared with raw medicinal powder (i.e., powder directly pulverized medicinal materials), extracts have significant advantages in terms of component purity, potency, and absorption rate. *Bifidobacterium animalis* subsp. *lactamase* is a Gram-positive anaerobic bacterium belonging to the genus *Bifidobacterium*. It has a short, curved rod-like or bifidoidal rod-like morphology, is non-motile, and does not form spores. This bacterium is a strictly anaerobic chemoheterotrophic microorganism, capable of producing acid from sucrose, lactose, and maltose, but exhibits only weak hydrolysis of milk. It requires multiple vitamins for growth and is negative for oxidases and catalases.

[0004] Existing technologies include studies on using traditional Chinese medicine extracts or Bifidobacterium lactis subsp. animalis to lower serum uric acid levels. However, the effects of using either alone are limited, and the existing Bifidobacterium lactis subsp. animalis has a single function. Further research is needed to find Bifidobacterium lactis subsp. animalis with diverse functions and excellent effects. Summary of the Invention

[0005] Purpose of the invention: In view of the problems existing in the prior art, the present invention provides a strain of Bifidobacterium lactis subspecies KW365 that can be fermented into food and medicine to lower uric acid and its application.

[0006] Technical solution: To achieve the above-mentioned objectives, the present invention adopts the following technical solution: In a first aspect, the present invention provides a strain of Bifidobacterium animalis subsp. lactis that can be fermented to lower uric acid in food and medicine. The Bifidobacterium animalis subsp. lactis is Bifidobacterium animalis subsp. lactis KW365, which is deposited at the China Center for Type Culture Collection on May 28, 2024, with accession number CCTCC NO: M 20241082.

[0007] In a second aspect, the present invention provides a composition comprising Bifidobacterium lactis subsp. KW365 as described in claim 1.

[0008] As one embodiment, the composition is a lyophilized powder containing Bifidobacterium animalis subsp. lactis KW365; preferably, the activity of Bifidobacterium animalis subsp. lactis KW365 in the lyophilized powder is 1.0 × 10⁻⁶. 10 ~1.0×10 12 CFU / g.

[0009] Thirdly, the present invention provides a fermentation product of a traditional Chinese medicine extract obtained by fermenting Bifidobacterium lactis subsp. KW365, wherein the traditional Chinese medicine extract is obtained by water extraction of kudzu root, dandelion and licorice separately and then combining the extracts, or by water extraction of the three together.

[0010] As a specific implementation plan: The conditions for water extraction are: a material-to-liquid ratio of 1:(8-12), and soaking at 50-80℃ for 3-5 hours; The fermented herbal extract culture medium has the following composition: peptone 8-12.0 g / L, beef extract 4.0-6.0 g / L, yeast extract 3.0-5.0 g / L, glucose 1.5-3.5 g / L, dipotassium hydrogen phosphate 1.0-3.0 g / L, triamine citrate 1.0-3.0 g / L, sodium acetate 4.0-6.0 g / L, magnesium sulfate 0.1-0.3 g / L, manganese sulfate 0.02-0.08 g / L, Tween 80 0.5-1.5 g / L, agar 14.0-16.0 g / L, calcium carbonate 1.0-2.0 g / L, with the herbal extract serving as the dissolution solution for the culture medium components. The fermentation conditions are as follows: Bifidobacterium animalis subsp. lactis KW365 is activated and inoculated into the above culture medium at an inoculation rate of 2-8%, and cultured at 35-40℃ for 10-30 hours.

[0011] Fourthly, the present invention provides the application of the aforementioned Bifidobacterium lactis subsp. KW365, the aforementioned composition, and the aforementioned fermentation product in the preparation of antibacterial agents; preferably, the antibacterial agent is capable of inhibiting Enterobacter cloacae, Streptococcus pharyngitis, and / or Vibrio parahaemolyticus.

[0012] Fifthly, the present invention provides the application of the aforementioned Bifidobacterium lactis subsp. KW365, the aforementioned composition, and the aforementioned fermentation product in the preparation of a product that kills poliovirus type 1.

[0013] Sixthly, the present invention provides a fermentation product of a traditional Chinese medicine extract obtained by fermenting Bifidobacterium lactis subsp. KW365, wherein the traditional Chinese medicine extract is obtained by water extraction of each of the following herbs: Coix seed, Eucommia leaf, Poria cocos, Pueraria lobata, Dioscorea opposita, Taraxacum mongolicum, Glycyrrhiza uralensis, and Hippophae rhamnoides, followed by combining the extracts, or by water extraction of the following herbs.

[0014] As a specific implementation plan: The raw materials for the herbal extract are as follows by weight: Job's tears 2-11 parts, Eucommia ulmoides leaves 3-9 parts, Poria cocos 1-7 parts, Pueraria lobata 1-7 parts 1-7 parts yam, 0.5-4 parts dandelion, 0.5-4 parts licorice, 0.5-4 parts sea buckthorn; Preferably, the raw materials of the herbal extract are in the following weight proportions: Job's tears 7 parts, Eucommia ulmoides leaves 5 parts, Poria cocos 3 parts, Pueraria lobata 3 parts 3 parts yam, 1 part dandelion, 1 part licorice, 1 part sea buckthorn; The conditions for water extraction are: a material-to-liquid ratio of 1:(8-12), heating to 35-95℃ and soaking for 1-6 hours; preferably, a material-to-liquid ratio of 1:10, heating to 65℃ and soaking for 3 hours; The conditions for fermenting the herbal extract are as follows: filter the herbal extract, add 0.6-1.0% by weight of enzymatically hydrolyzed whey protein powder or soy peptone, sterilize, and inoculate with Bifidobacterium animalis subsp. lactis KW365 at an inoculation rate of 2-8% for fermentation for 30-40 hours; preferably, add 0.8% by weight of enzymatically hydrolyzed whey protein powder, with an inoculation rate of 5%, and ferment for 36 hours.

[0015] In a seventh aspect, the present invention provides the use of the aforementioned Bifidobacterium lactis subsp. KW365, the aforementioned composition, and the aforementioned fermentation product in the preparation of products that lower serum uric acid.

[0016] Eighthly, the present invention provides the application of the aforementioned Bifidobacterium lactis subsp. KW365, the aforementioned composition, and the aforementioned fermentation product in the preparation of products for protecting against kidney injury.

[0017] Beneficial Effects: Compared with existing technologies, this invention provides a novel *Bifidobacterium animalis* subspecies KW365, which exhibits good tolerance to traditional Chinese medicine. Both the strain itself and the fermentation products prepared by fermenting specific traditional Chinese medicines using KW365 demonstrate significant antibacterial and antiviral effects, particularly effectively inhibiting *Enterobacter cloacae*, *Streptococcus pharyngitis*, and / or *Vibrio parahaemolyticus*, and killing poliovirus type 1. Furthermore, by fermenting specific food-medicine homologous ingredients using KW365, the strain itself and the fermentation products effectively inhibit xanthine oxidase (XOD) and adenosine deaminase (ADA) activities, thereby effectively reducing serum uric acid levels, alleviating hyperuricemia, and protecting against kidney damage. The fermentation products show particularly superior effects, indicating a significant synergistic effect between KW365 and specific traditional Chinese medicines. Attached Figure Description

[0018] Figure 1 Gram staining image of Bifidobacterium lactis strain KW365.

[0019] Figure 2 The changes in serum uric acid (SUA) levels in mice from different groups were compared.

[0020] Figure 3 Comparison of blood urea nitrogen (BUN) concentrations in mice from different groups.

[0021] Figure 4 Comparison of serum creatinine (CRE) concentrations in mice from different groups.

[0022] Figure 5 The effects of each group on serum adenosine deaminase (ADA) in mice.

[0023] Figure 6 The effects of each group on serum xanthine oxidase (XOD) in mice were investigated. Detailed Implementation

[0024] The technical solution of the present invention will be further described in detail below with reference to the accompanying drawings and embodiments.

[0025] Example 1: Screening of highly active lactic acid bacteria strains and construction of a strain candidate library To obtain beneficial lactic acid bacteria strains for human use, samples were collected from healthy individuals, traditional yogurt starter cultures, and sauerkraut as target bacterial sources. This aimed to obtain healthy reproductive tract flora in one step, which could then be used to intervene in disordered reproductive tract flora. Dominant strains of healthy reproductive tract flora were screened. After thawing, samples were serially diluted 10-fold. Each serial dilution was then plated on MRS agar medium, with four dips per gradient and five parallel dilutions per sample. The plates were incubated in anaerobic and 5% oxygen environments at 35-37℃. After 48 hours of incubation, independent, round, and well-grown colonies from each gradient were selected. MALDI-TOF mass spectrometry was used for rapid preliminary identification of the selected colonies. Known pathogenic and opportunistic pathogens were discarded. Probiotics or second-generation probiotics were inoculated into 5 ml of MRS liquid medium for large-scale culture. After 48 hours of incubation at 35-37℃, the OD of the fermentation broth was measured. 600 The bacterial count in the fermentation broth was observed by comparison with microscopic observation. For well-fermenting strains, 2 ml of the culture was added to 2 ml of 30%-40% glycerol solution, pre-cooled, and then frozen at -80°C for further isolation, purification, and screening. Samples with poor fermentation or no obvious bacterial amplification were added to 5 ml of fresh MRS liquid medium and cultured again for 48 hours. The OD of the fermentation broth was then measured. 600 The bacterial count in the fermentation broth was observed by comparison with a microscope. 2 ml of the well-fermented strains were added to 2 ml of 30%-40% glycerol solution, pre-cooled, and then frozen at -80℃ for further isolation, purification, and screening. Strains that did not ferment were discarded. In this way, strains with high activity and easy culture were screened. A total of 588 strains were screened according to this process, which served as a preliminary strain candidate library.

[0026] Example 2 Screening of uric acid-lowering strains Some of the strains screened in Example 1 were isolated and purified. The purified strains were then inoculated into MRS medium for proliferation culture, and OD was adjusted. 600The value is 1.2 for later use. A screening medium for strains capable of decomposing uric acid was prepared, using MRS medium as a reference, but with glucose removed and uric acid replaced as the carbon source. The composition of the MRS-C screening medium was: peptone 10.0 g / L, beef extract 5.0 g / L, yeast extract 4.0 g / L, uric acid 10 g / L, dipotassium hydrogen phosphate 2.0 g / L, triamine citrate 2.0 g / L, sodium acetate 5.0 g / L, magnesium sulfate 0.2 g / L, manganese sulfate 0.05 g / L, and Tween 80 1.0 g / L. An additional 15.0 g / L agar and 1.5 g / L calcium carbonate were added to the agar solid medium. Uric acid plates were prepared using Oxford cups. 200 μL of bacterial suspension was inoculated into each well of the Oxford cup and incubated at 37°C for 24-36 hours. The size of the calcium dissolution zone was observed; a larger calcium dissolution zone indicated better bacterial growth on the MRS-C selection medium, and these strains were selected as target strains. Finally, 15 strains exhibiting larger calcium dissolution zones on the uric acid-degrading selective medium were chosen as backup strains.

[0027] Example 3 Screening of strains resistant to / fermentable traditional Chinese medicine Some medicinal and edible components or traditional Chinese medicines have varying degrees of antibacterial ability. To obtain strains that have a certain tolerance to the antibacterial components of traditional Chinese medicines or can ferment them, kudzu root, dandelion, and licorice were selected for screening and evaluation. Kudzu root, dandelion, and licorice were ultra-finely pulverized, and 1g of each was added to 10ml of pure water. The mixture was heated to 65℃ and soaked for 4 hours, then cooled to room temperature and filtered. The pulverized mixture was then mixed in a 1:1:1 ratio to obtain the mixed extract of traditional Chinese medicines (MED-3). The MRS-resistant screening medium for traditional Chinese medicines (MRS-MED-3) was prepared by reducing the glucose in the MRS by 50% and using the mixed extract of traditional Chinese medicines as the solution for the medium components. The composition of the screening medium MRS-MED-3 was as follows: peptone 10.0 g / L, beef extract 5.0 g / L, yeast extract 4.0 g / L, glucose 2.5 g / L, dipotassium hydrogen phosphate 2.0 g / L, triamine citrate 2.0 g / L, sodium acetate 5.0 g / L, magnesium sulfate 0.2 g / L, manganese sulfate 0.05 g / L, and Tween 80 1.0 g / L. An additional 15.0 g / L agar and 1.5 g / L calcium carbonate were added to the MRS-MED-3 liquid medium. Oxford cups were used to prepare MRS screening plates for resistance to traditional Chinese medicine. Bacterial suspensions (OD) of the 15 strains screened in Example 2 were collected from each strain. 600 (Value 1.2) 200 μL was inoculated into the wells of an Oxford cup and incubated at 37℃ for 24-36 h. The size of the calcium dissolution zone was observed. A larger calcium dissolution zone indicates that the bacteria reproduce well on MRS-MED-3 selection medium and have the characteristics of tolerance to traditional Chinese medicine. Simultaneously, the strain with a larger calcium dissolution zone was inoculated at 5% into MRS-MED-3 liquid medium and incubated at 37℃ for 24 h. The OD of the fermentation broth was measured. 600The bacterial morphology was observed under a microscope, and the bacterial count in the fermentation broth was compared. The strain that fermented well could be used to ferment Chinese herbal medicines with antibacterial properties. The target strain was finally numbered KW365.

[0028] Example 4: Preservation, identification, and archiving of strains Based on the evaluation results of the strain screening process in Examples 1-3, strains that possess high activity, can metabolize uric acid, have a certain tolerance to the antibacterial components of traditional Chinese medicine, and can ferment traditional Chinese medicine were identified and collected.

[0029] (1) Preservation of strains After streaking the selected KW365 strain onto MRS agar medium for 48 hours, single colonies were selected and placed in 10 ml of liquid culture medium. This medium was then incubated at 37°C for 12-16 hours, and the OD of the culture medium was measured. 600 When the value is ≥0.6, add glycerol solution at a volume ratio of 1:1 to the culture medium. The concentration of the glycerol solution is 30%-40%. Mix well by pipetting and dispensing into 2ml sterile cryovials. Pre-cool at 4℃ for 2h, then pre-freeze at -20℃ for 4h, and finally transfer to a -80℃ freezer or liquid nitrogen for storage as the 0th generation original strain.

[0030] (2) Further morphological identification The bacteria on agar medium appear as round, rod-shaped colonies with neat edges, milky white color, a moist and smooth surface, and a raised center. Microscopic observation of the purified bacterial solution reveals that they are Gram-positive, rod-shaped, with some having forked or curved ends, lacking flagella and spores.

[0031] (3) Gram staining Gram staining result is Gram positive, such as Figure 1 As shown.

[0032] (4) Molecular biological identification of 16S rRNA The 16S rDNA gene sequence of strain KW365 was amplified and sequenced using published universal 16S primers (primer sequences: 8F: 5'-AGAFTTTGATCCTGGCTCA-3'; 1510R: 5'-GGTTACCTTGTTACGACTT-3'). The nucleotide sequence of the 16S rDNA of strain KW365 is sequence 1 in the sequence listing. After 16S rDNA gene alignment, the similarity rate with Bifidobacterium animalis subsp. lactis in Genebank reached 99%. Combined with microbial systematic identification, strain KW365 was identified as a subsp. lactis of Bifidobacterium animalis and named Bifidobacterium animalis subsp. lactis KW365.

[0033] Bifidobacterium animalis subsp. lactis KW365, identified by biological analysis, was deposited at the China Center for Type Culture Collection (CCTCC) on May 28, 2024, with accession number CCTCC M 20241082, at Wuhan University, Wuhan, Hubei Province, People's Republic of China.

[0034] Example 5: In vitro antibacterial experiment To test the in vitro antibacterial activity of *Bifidobacterium lactis* subsp. KW365, *Bifidobacterium lactis* HN019 was selected as the control strain. Both strains were inoculated into 50 ml of MRS liquid medium for activation and cultured at 37°C for 16 h. The fermentation broth was then centrifuged at 10000 rpm for 10 min, and the supernatant and bacterial cells were collected separately. The bacterial cells were washed twice with 0.9% sterile saline, and then OD was prepared using 0.9% sterile saline. 600 The bacterial suspension is 0.6 g / L and stored at 4°C for future use.

[0035] Antibacterial activity was determined using the agar diffusion method. A double-layer plate was used, with the bottom layer of water agar culture inverted, and sterile Oxford cups placed on solidified water agar. Enterobacter cloacae ATCC23355, Streptococcus pharyngitis ATCC 9895, and Vibrio parahaemolyticus ATCC17802 were selected as test strains for in vitro antibacterial experiments. Test strains activated to the logarithmic phase were diluted with sterile physiological saline to OD0.05. 600For a bacterial suspension with a concentration of 0.5-0.8, 1 mL of the diluted suspension was added to 50 mL of MRS agar medium at approximately 40°C and quickly mixed thoroughly. 30 mL of medium containing indicator bacteria was then poured into a petri dish containing Oxford cups. After solidification, the Oxford cups were removed. The pH of the MRS culture medium was adjusted with lactic acid to match that of the supernatant as a blank control. A 0.05 mg / mL erythromycin solution was used as a positive control. 200 μL each of the KW365 and HN019 bacterial suspensions and supernatants were used for an inhibition test, and the diameter of the inhibition zone was measured.

[0036] The results are shown in Table 1. Compared with the positive control group and the blank control group, the suspension and supernatant of Bifidobacterium animalis subsp. lactis KW365 showed significant inhibitory effects on Enterobacter cloacae, Streptococcus pharyngitis, and Vibrio parahaemolyticus. Moreover, the inhibitory ability of the suspension of Bifidobacterium animalis subsp. lactis KW365 against the three pathogenic bacteria was significantly better than that of Bifidobacterium lactis HN019. It can be seen that the fermentation metabolites of Bifidobacterium animalis subsp. lactis KW365 contain antibacterial components, and the bacterial cells can still inhibit pathogenic bacteria. It can be used in intervention preparations to inhibit pathogenic bacteria and regulate the flora.

[0037] Table 1 Evaluation of the antibacterial activity of Bifidobacterium animalis subsp. lactis KW365

[0038] Note: "-" indicates no obvious inhibition zone appears, "+" indicates the diameter of the inhibition zone is between 3mm and 5mm, "++" indicates the diameter of the inhibition zone is between 5mm and 10mm, and "+++" indicates the diameter of the inhibition zone is greater than 10mm.

[0039] Example 6 Evaluation of the antibacterial activity of fermented traditional Chinese medicine by Bifidobacterium animalis subsp. lactis KW365 Following the method in Example 3, KW365 was activated and inoculated at 5% into MRS-MED-3 liquid medium. After incubation at 37°C for 16 hours, the fermentation broth was centrifuged at 10000 rpm for 10 minutes, and the supernatant and bacterial cells were collected separately. The bacterial cells were washed twice with 0.9% sterile saline, and then OD was prepared using 0.9% sterile saline. 600The bacterial suspension was prepared at 0.6% and stored at 4°C for future use. The pH of the MRS culture medium was adjusted with lactic acid to match that of the supernatant as a blank control, and a 0.05 mg / mL erythromycin solution was used as a positive control. The antibacterial activity of *Bifidobacterium lactis* subsp. *majorheicum* KW365 fermented with traditional Chinese medicine was evaluated according to the antibacterial evaluation test method in Example 5, and the results are shown in Table 2. Combined with the evaluation results of Example 5, the fermentation broth obtained by fermenting the mixed extract of traditional Chinese medicine with *Bifidobacterium lactis* subsp. *majorheicum* KW365 significantly improved the inhibitory activity against *Enterobacter cloacae* and *Streptococcus pharyngitis*, and its antibacterial activity was superior to that of the unfermented mixed extract of traditional Chinese medicine (MED-3), indicating that fermentation of traditional Chinese medicine with *Bifidobacterium lactis* subsp. *majorheicum* KW365 can enhance the antibacterial activity of traditional Chinese medicine.

[0040] Table 2 Evaluation of the antibacterial effect of fermented traditional Chinese medicine containing Bifidobacterium animalis subsp. lactis KW365

[0041] Note: "-" indicates no obvious inhibition zone appears, "+" indicates the diameter of the inhibition zone is between 3mm and 5mm, "++" indicates the diameter of the inhibition zone is between 5mm and 10mm, and "+++" indicates the diameter of the inhibition zone is greater than 10mm.

[0042] Example 7: Virus-killing test of fermentation broth of traditional Chinese medicine According to the virus inactivation test method in the "Disinfection Technical Specifications", poliovirus type 1 (PV-I) vaccine strain was used as the experimental virus strain, and VERO cell line was used as the test cell. Virus inactivation experiments were performed on the MRS-MED-3 fermentation supernatant from Example 3. The virus content in the cell culture was detected using indirect immunofluorescence assay. The original fermentation broth was used as the test concentration, and MRS-MED-3 medium was used as the control. The Reed-Muench method was used to calculate the cellular half-maximal infectious dose (TCID) of the virus in each sample. 50 As shown in Table 3, the evaluation results of the virus-killing ability show that the fermentation broth produced by Bifidobacterium animalis subsp. lactis KW365 through the fermentation of traditional Chinese medicine exhibits excellent virus-killing ability.

[0043] Table 3. Killing effect of culture supernatant of various Chinese herbal medicine extracts on poliovirus (treatment time 5 min).

[0044] Note: "*" indicates that the absolute 100% kill rate was not achieved; the result is rounded to two decimal places and considered 100%. Example 8: Preparation of freeze-dried powder with high storage stability (1) Culture medium preparation.

[0045] The following is a culture medium preparation scheme for industrial-scale high-density fermentation of Bifidobacterium animalis subsp. lactis KW365: 8 g / L soybean peptone, 8 g / L yeast extract, 10 g / L hydrolyzed whey protein, 22 g / L anhydrous glucose, 3 g / L tomato puree (25% solids), 2 g / L dipotassium hydrogen phosphate, 2 g / L diamine hydrogen citrate, 5 g / L sodium acetate, 0.5 g / L manganese sulfate, 0.5 g / L L-cysteine ​​sulfate, and 1 ml / L Tween 80. All raw materials are food-grade.

[0046] (2) Liquid culture After preparing the culture medium, adjust the pH to 6.5, sterilize at 115℃ for 30 minutes, and cool to 38℃-39℃. Inoculate the fermenter at a rate of 3%-10%, preferably 8% in this example. The culture temperature is 35℃-37℃, and the pH is 6.2-6.5. During fermentation, the fermenter is sealed and pressure is maintained using nitrogen. Stir slowly. Continue culturing for 12-20 hours. At 12 hours, the viable cell count in the fermentation broth is measured to be 6.8 × 10⁻⁶. 8 The CFU / mL concentration was measured at 20 h, and the viable cell count in the fermentation broth was 2.5 × 10⁻⁶. 9 The CFU / mL level indicates a relatively high level of viable fermentation cells, meeting the requirements for high-density industrial fermentation production.

[0047] (3) Centrifugation To avoid prolonged fermentation leading to bacterial aging and affecting the activity of the freeze-dried powder, the viable cell count should reach 10^6 microorganisms after 12-20 hours of cultivation. 8 CFU / mL-10 9 CFU / mL, based on OD 600 After the growth curve reached the plateau phase, the fermentation broth was quickly cooled to below 15°C and centrifuged at 5000 rpm using a tubular centrifuge to harvest the mycelium sludge.

[0048] (4) Emulsification and freeze-drying of protective agents After 16 hours of fermentation, the centrifuged and harvested sludge was diluted to a moisture content of 60%-80%, and an equal volume of sludge preservative was added for emulsification. The sludge preservative consisted of: 70% water, 10% skim milk powder, 5% sucrose, 5% mannitol, 1% Tween-80, 3% betaine, 0.5% monosodium glutamate, and 5% soluble starch. The dried material was characterized by a moisture content of less than 5% and a water activity of 0.08-0.20 aw.

[0049] The freeze-dried bacterial powder had an activity of 6.5 × 10⁻⁶. 11 With a CFU / g, it meets the high-yield requirements and can be used as a food additive for lactic acid bacteria powder or for the preparation of microecological intervention preparations.

[0050] Example 9 Preparation of foriiX, a fermentation product that is both food and medicine Option 1: 7 parts of Job's tears (X1), 5 parts of Eucommia ulmoides leaves (X2), 3 parts of Poria cocos (X3), 3 parts of kudzu root (X4), 3 parts of yam (X5), 1 part of dandelion (X6), 1 part of licorice root (X7), and 1 part of sea buckthorn (X8) are ultra-finely pulverized and then mixed with 10 times their dry weight of water. The mixture is soaked at 65℃ for 3 hours. After filtering the extract, 0.8% enzymatically hydrolyzed whey protein powder is added. The mixture is then sterilized at 115℃ and inoculated with 5% Bifidobacterium animalis subsp. lactis KW365 for fermentation for 36 hours. The fermentation broth is then filtered and concentrated using a nano-ceramic membrane. The concentrate is then freeze-dried under vacuum to maintain the effective activity of the fermentation product, resulting in the medicinal and edible fermentation product foriiX.

[0051] The following methods can be selected depending on the amount of raw materials used and the different processes: Option 2: Combine 2 parts of Job's tears (X1), 3 parts of Eucommia ulmoides leaves (X2), 1 part of Poria cocos (X3), 1 part of kudzu root (X4), 1 part of yam (X5), 0.5 parts of dandelion (X6), 0.5 parts of licorice root (X7), and 0.5 parts of sea buckthorn (X8). After ultra-fine grinding, add 10 times the amount of water by dry weight and soak at 35℃ for 3 hours. Filter the extract and add 0.8% soybean peptone. Sterilize at 115℃ and inoculate with 5% Bifidobacterium animalis subsp. lactis KW365 for fermentation for 36 hours. Filter and concentrate the fermentation broth using a nano-ceramic membrane. Freeze-dry the concentrate under vacuum to maintain the effective activity of the fermentation product, resulting in the medicinal and edible fermentation product forii2.

[0052] Option 3: 11 parts of Job's tears (X1), 9 parts of Eucommia ulmoides leaves (X2), 7 parts of Poria cocos (X3), 7 parts of kudzu root (X4), 7 parts of yam (X5), 4 parts of dandelion (X6), 4 parts of licorice root (X7), and 3 parts of sea buckthorn (X8) were ultra-finely pulverized and then mixed with 10 times their dry weight of water. The mixture was soaked at 95℃ for 3 hours. After filtering the extract, 0.8% soybean peptone was added. The mixture was then sterilized at 115℃ and inoculated with 5% Bifidobacterium animalis subsp. lactis KW365 for fermentation for 36 hours. The fermentation broth was filtered and concentrated using a nano-ceramic membrane. The concentrate was then freeze-dried under vacuum to maintain the effective activity of the fermentation product, resulting in the medicinal and edible fermentation product forii3.

[0053] Similar to other food and medicine fermentation, the resulting freeze-dried fermentation metabolites can be used to prepare powders, tablets, liquids and other dosage forms.

[0054] Example 10: Interventional effect of Bifidobacterium animalis subsp. lactis KW365 on hyperuricemia. Fifty-six male Kunming mice (6 weeks old, 25±2g) were randomly divided into seven groups (n=8 / group): negative control group (NC), hyperuricemia group (HUA), KW365 intervention group (KW365), foriiX intervention group (foriiX), forii2 intervention group (forii2), forii3 intervention group (forii3), and foriiX control group (foriiX-NC). Hyperuricemia was induced in the HUA, intervention, and intervention control groups by suspending potassium oxychloride and adenine in 0.5% sodium carboxymethyl cellulose solution and administering 0.3 ml of potassium oxychloride (250 mg / kg) and adenine (75 mg / kg) solution daily via tube feeding for 14 days. Mice in the NC group were given an equal volume of 0.5% sodium carboxymethyl cellulose solution for 14 days. Mice in the KW365 intervention group were given 0.3 ml of PBS bacterial solution (containing 1.0 × 10⁻⁶ Bifidobacterium lactis subsp. KW365) 4 hours after being fed potassium oxychloride and adenine. 9 Mice in the CFU group, foriiX, forii2, and forii3 intervention groups were given 0.3 ml of their respective fermentation broth (lyophilized powder prepared according to Example 9, 1 g dissolved in 5 ml of sterile PBS). ForriX-NC was given 0.3 ml of a medicinal and edible extract (lyophilized powder prepared according to Scheme 1 of Example 9, sterilized without KW365 fermentation medium, 1 g dissolved in 5 ml of sterile PBS). Mice in the NC and HUA groups were given an equal volume of sterile PBS. After 14 days of continuous intervention, mice were sacrificed, weighed, and blood was collected from the ocular vein to detect serum uric acid (SUA) concentration. The results are as follows: Figure 2 As shown.

[0055] Compared with the NC group mice, the HUA group mice showed significantly increased serum uric acid (SUA) concentration and poorer health, indicating the success of the HUA mouse model. Oral administration of Bifidobacterium animalis subsp. lactis KW365 significantly reduced serum uric acid (SUA) levels, thus confirming that Bifidobacterium animalis subsp. lactis KW365 can lower serum uric acid (SUA). The fermentation product foriiX, derived from Bifidobacterium animalis subsp. lactis KW365 fermented with food-grade ingredients, showed a significantly better effect on lowering serum uric acid in mice than both Bifidobacterium animalis subsp. lactis KW365 and the foriiX-NC control group. This indicates that fermentation of food-grade ingredients with Bifidobacterium animalis subsp. lactis KW365 can effectively enhance the effect of lowering serum uric acid, demonstrating a synergistic effect between the two in reducing serum uric acid. Meanwhile, the effects of fermentation products of medicinal and edible ingredients with different formulations on lowering serum uric acid were compared. The effect of foriiX on lowering serum uric acid was better than that of forii2 and forii3. Therefore, the preparation scheme of foriiX (2 parts of coix seed (X1), 3 parts of eucommia leaf (X2), 1 part of poria cocos (X3), 1 part of kudzu root (X4), 1 part of yam (X5), 0.5 parts of dandelion (X6), 0.5 parts of licorice (X7), and 0.5 parts of sea buckthorn (X8), after ultra-fine grinding, add 10 times the amount of water according to dry weight, soak at 35℃ for 3 hours, filter the extract, add 0.8% soybean peptone, sterilize at 115℃, and then inoculate with Bifidobacterium animalis subsp. lactis KW365 for fermentation for 36 hours) was selected as the optimal preparation scheme for lowering serum uric acid.

[0056] Example 11: Protective effect of food-medicine homologous fermentation products on kidney damage induced by HUA. Blood urea nitrogen (BUN) and serum creatinine (CRE) are important indicators of kidney injury. The concentrations of BUN and CRE in the serum of mice collected in Example 10 were measured, and the results are as follows: Figure 3 , Figure 4 As shown in the figure. The results showed that compared with NC mice, the levels of blood urea nitrogen (BUN) and serum creatinine (CRE) in the HUA group mice were significantly increased. Oral administration of Bifidobacterium lactis subsp. KW365 or foriiX effectively reversed the increase in blood urea nitrogen (BUN) and serum creatinine (CRE) levels induced by HUA, and the intervention effect of foriiX was better. This indicates that Bifidobacterium lactis subsp. KW365 has a protective effect against kidney injury, and the medicinal and food homologous fermentation product foriiX fermented by Bifidobacterium lactis subsp. KW365 has a better effect. The two have a synergistic effect.

[0057] Example 12: The alleviating effect of Bifidobacterium animalis subsp. lactis KW365 and its ferment foriiX on hyperuricemia. Since uric acid (UA) is mainly synthesized in the liver, xanthine oxidase (XOD) and adenosine deaminase (ADA) are two essential enzymes for uric acid synthesis in the liver. The activities of xanthine oxidase (XOD) and adenosine deaminase (ADA) were measured in the serum of mice collected in Example 10. The results are as follows: Figure 5 , Figure 6 As shown, compared with the control group (NC) mice, the activities of xanthine oxidase (XOD) and adenosine deaminase (ADA) were significantly increased in hyperuricemic (HUA) mice. Intervention with foriiX and KW365 significantly inhibited the activities of adenosine deaminase (ADA) and xanthine oxidase (XOD). Furthermore, the inhibitory effects of foriiX-NC, forii2, and forii3 in the experimental control group were also observed. This indicates that oral administration of Bifidobacterium animalis subsp. lactis KW365 can effectively reduce serum uric acid (SUA) levels and alleviate symptoms of hyperuricemia (HUA). The medicinal and edible fermentation product foriiX, fermented with Bifidobacterium animalis subsp. lactis KW365, has a better inhibitory effect on xanthine oxidase (XOD) and adenosine deaminase (ADA) activities, exhibiting a significant synergistic effect.

[0058] The embodiments of the present invention have been described in detail above with reference to specific examples. However, the present invention is not limited to the above embodiments. Within the scope of knowledge possessed by those skilled in the art, various changes can be made without departing from the spirit of the present invention.

Claims

1. An animal Bifidobacterium lactis strain capable of fermenting a medicinal and edible material to reduce uric acid, characterized in that, The animal Bifidobacterium lactis is Bifidobacterium animalis subsp. lactis KW365, which is preserved in China Center for Type Culture Collection on May 28, 2024, and the preservation number is CCTCC NO: M 20241082.

2. A composition characterized in that, The composition comprises the animal Bifidobacterium lactis KW365 of claim 1.

3. A fermentation product of a Chinese herbal medicine extract fermented with the Bifidobacterium animalis ssp. lactis KW365 according to claim 1, characterized by, The Chinese herbal medicine extract is obtained by water extraction of Pueraria lobata, Taraxacum mongolicum, and Glycyrrhiza uralensis respectively, or by water extraction of a mixture of the three.

4. The fermentation product of claim 3, wherein, The water extraction condition is that the solid-liquid ratio is 1: (8-12), and the soaking is performed at 50-80 DEG C for 3-5 h. The fermentation medium comprises the following components in the following proportions: 8-12.0 g / L of proteose peptone, 4.0-6.0 g / L of beef extract, 3.0-5.0 g / L of yeast extract, 1.5-3.5 g / L of glucose, 1.0-3.0 g / L of potassium phosphate dibasic, 1.0-3.0 g / L of triamine citrate, 4.0-6.0 g / L of sodium acetate, 0.1-0.3 g / L of magnesium sulfate, 0.02-0.08 g / L of manganese sulfate, 0.5-1.5 g / L of Tween 80, 14.0-16.0 g / L of agar, 1.0-2.0 g / L of calcium carbonate, and the Chinese herbal medicine extract as a medium component solution. The fermentation condition is that the activated animal Bifidobacterium lactis KW365 is inoculated into the above-mentioned medium at an inoculation amount of 2-8%, and the culture is performed at 35-40 DEG C for 10-30 h.

5. The animal Bifidobacterium lactis KW365 of claim 1, the composition of claim 2, or the fermentation product of claim 3 or 4 is used for preparing an antibacterial agent; preferably, the antibacterial agent can inhibit Enterobacter cloacae, Streptococcus anginosus, and / or Vibrio parahaemolyticus.

6. The animal Bifidobacterium lactis KW365 of claim 1, the composition of claim 2, or the fermentation product of claim 3 or 4 is used for preparing a product for killing poliovirus type 1.

7. A fermentation product of a Chinese herbal medicine extract fermented with the Bifidobacterium animalis ssp. lactis KW365 according to claim 1, characterized by, The Chinese herbal medicine extract is obtained by water extraction of Pueraria lobata, Taraxacum mongolicum, and Glycyrrhiza uralensis respectively, or by water extraction of a mixture of the three.

8. The fermentation product of claim 7, wherein, The weight parts of the raw materials of the Chinese herbal medicine extract are as follows: Pueraria lobata 2-11 parts, Eucommia ulmoides leaf 3-9 parts, Poria cocos 1-7 parts, Pueraria lobata 1-7 parts, Pueraria lobata 2-11 parts, Eucommia ulmoides leaf 3-9 parts, Poria cocos 1-7 parts, Pueraria lobata 1-7 parts, Pueraria lobata 2-11 parts, Eucommia ulmoides leaf 3-9 parts, Poria cocos 1-7 parts, Pueraria lobata 1-7 parts, Pueraria lobata 2-11 parts, Eucommia ulmoides leaf 3-9 parts, Poria cocos 1-7 parts, Pueraria lobata 1-7 parts, The water extraction condition is that the solid-liquid ratio is 1: (8-12), and the soaking is performed at 50-80 DEG C for 3-5 h. The fermentation condition is that the activated animal Bifidobacterium lactis KW365 is inoculated into the above-mentioned medium at an inoculation amount of 2-8%, and the culture is performed at 35-40 DEG C for 10-30 h. The fermentation of the Chinese herbal medicine extract is as follows: the Chinese herbal medicine extract is filtered, 0.6-1.0% of enzyme-degraded whey protein powder or soybean peptone by weight is added, and then the mixture is sterilized and inoculated with Bifidobacterium animalis lactis KW365 at an inoculation amount of 2-8% for fermentation, and the fermentation time is 30-40 h; preferably, 0.8% of enzyme-degraded whey protein powder by weight is added, the inoculation amount is 5%, and the fermentation time is 36 h.

9. Use of the Bifidobacterium animalis lactis KW365 of claim 1, the composition of claim 2, or the fermentation product of claim 7 or 8 in the preparation of a product for reducing serum uric acid.

10. Use of the Bifidobacterium animalis lactis KW365 of claim 1, the composition of claim 2, or the fermentation product of claim 7 or 8 in the preparation of a product for protecting kidney injury.

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